Formation of Cyclopropanone during Cytochrome P450-Catalyzed N-Dealkylation of a Cyclopropylamine

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• 作者：Christopher L. Shaffer ; Shawn Harriman ; Yakov M. Koen ; and Robert P. Hanzlik
• 刊名：Journal of the American Chemical Society
• 出版年：2002
• 出版时间：July 17, 2002
• 年：2002
• 卷：124
• 期：28
• 页码：8268 - 8274
• 全文大小：83K
• 年卷期：v.124,no.28(July 17, 2002)
• ISSN：1520-5126

文摘

The role of single electron transfer (SET) in P450-catalyzed N-dealkylation reactions has beenstudied using the probe substrates N-cyclopropyl-N-methylaniline (2a) and N-(1'-methylcyclopropyl)-N-methylaniline (2b). In earlier work, we showed that SET oxidation of 2a by horseadish peroxidase leadsexclusively to products arising via fragmentation of the cyclopropane ring [Shaffer, C. L.; Morton, M. D.;Hanzlik, R. P. J. Am. Chem. Soc. 2001, 123, 8502-8508]. In the present study, we found that livermicrosomes from phenobarbital pretreated rats (which contain CYP2B1 as the predominant isozyme) oxidize[1'-¹³C, 1'-¹⁴C]-2a efficiently (80% consumption in 90 min). Disappearance of 2a follows first-order kineticsthroughout, indicating a lack of P450 inactivation by 2a. HPLC examination of incubation mixtures revealedthree UV-absorbing metabolites: N-methylaniline (4), N-cyclopropylaniline (6a), and a metabolite (M1)tentatively identified as p-hydroxy-2a, in a 2:5:2 mole ratio, respectively. 2,4-Dinitrophenylhydrazine trappingindicated formation of formaldehyde equimolar with 6a; 3-hydroxypropionaldehyde and acrolein were notdetected. Examination of incubations of 2a by ¹³C NMR revealed four ¹³C-enriched signals, three of whichwere identified by comparison to authentic standards as N-cyclopropylaniline (6a, 33.6 ppm), cyclopropanonehydrate (11, 79.2 ppm), and propionic acid (12, 179.9 ppm); the fourth signal (42.2 ppm) was tentativelydetermined to be p-hydroxy-2a. Incubation of 2a with purified reconstituted CYP2B1 also afforded 4, 6a,and M1 in a 2:5:2 mole ratio (by HPLC), indicating that all metabolites are formed at a single active site.Incubation of 2b with PB microsomes resulted in p-hydroxylation and N-demethylation only; no loss orring-opening of the cyclopropyl group occurred. These results effectively rule out the participation of a SETmechanism in the P450-catalyzed N-dealkylation of cyclopropylamines 2a and 2b, and argue strongly forthe N-dealkylation of 2a via a carbinolamine intermediate formed by a conventional C-hydroxylationmechanism.